Accessory subunits are integral for assembly and function of human mitochondrial complex I

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Biology

Background:

• Complex I (NADH:ubiquinone oxidoreductase) is a large, essential enzyme in the mitochondrial respiratory chain, vital for ATP synthesis.
• Complex I dysfunction is implicated in mitochondrial diseases, Parkinson's disease, and aging, highlighting its clinical significance.
• While 14 core subunits are known, the roles of 31 human accessory subunits in Complex I assembly and function remain largely unknown.

Purpose of the Study:

• To investigate the necessity and function of the 31 accessory subunits of human Complex I.
• To determine the impact of accessory subunit loss on Complex I assembly, stability, and cellular viability.
• To elucidate the contribution of accessory subunits to the structural integrity and functional output of Complex I.

Main Methods:

• Generation of human knockout cell lines for each of the 31 accessory subunits using gene editing.
• Quantitative proteomic analysis of knockout cell lines to assess subunit stability and complex assembly.
• Investigation of specific module requirements for distal membrane arm assembly.

Main Results:

• 25 out of 31 accessory subunits were found to be strictly required for the assembly of a functional Complex I.
• One accessory subunit was identified as essential for cell viability.
• Loss of individual subunits destabilized other subunits within the same structural module, indicating interconnectedness.
• ATP5SL and DMAC1 were identified as crucial for the assembly of the distal membrane arm of Complex I.

Conclusions:

• Accessory subunits play a broad and critical role in the structure, assembly, and function of human Complex I.
• Gene editing coupled with proteomics provides a powerful approach to dissecting large multi-subunit complexes.
• Understanding the role of accessory subunits is crucial for comprehending Complex I dysfunction in disease and aging.